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Recent evidence suggests that metastasis is proceeded by premetastatic niche formation where bone marrow-derived cells (BMDC), particularly those of myeloid cells (monocytes and macrophages), form clusters preparing for the arrival of cancer cells in the metastasis-prone organs such as the lung. Many molecules involved in the premetastatic niche formation has been identified, some of which include vascular endothelial growth factor (VEGF), lysyl oxidase (LOX), and S100A8. Since VEGF, LOX, and S100A8 are downstream targets of hypoxia-inducible factor (HIF), a major transcription factor stabilized under hypoxic conditions, we investigated whether HIF-1 in myeloid cells would affect the premetastatic niche formation hence metastasis to the lung. To do this, we used our novel strain of myeloid specific Hif-1α knockout (KO) mice using human S100A8, an intracellular calcium-binding protein as the myeloid promoter. We first performed bone marrow (BM) transplantation from GFP-expressing myeloid specific Hif-1α KO (hereafter denoted as Hif-1α KO) mice or GFP-expressing littermate control mice to lethally irradiated wild-type (WT) mice. Two weeks later, these mice were subcutaneously implanted for Lewis lung carcinoma (LLC) primary tumors followed by 20 Gy chest irradiation to promote the premetastatic niche formation. Primary tumors were then removed and luciferase-expressing LLC cells were intravenously injected to induce the lung metastasis. We observed to our surprise that mice receiving Hif-1α KO BM had 10-fold increase in the lung metastasis, indicating that HIF-1α in myeloid cells is required to protect the lung from the metastasis. While the total number of GFP-expressing BMDC was similar, there were significantly decreased alveolar macrophages (GFP+F4/80+CD11c+CD11b-) in the irradiated lung of mice receiving BM of Hif-1α KO BM than the littermate control mice. Because alveolar macrophages are specialized immune cells involved in phagocytosis of numerous airborne antigens, we hypothesized that HIF-1α-deficient alveolar macrophages are deficient in phagocytosis and this may be responsible for the increase in the lung metastasis. Indeed, we observed that alveolar macrophages isolated from bronchioalveolar lavage fluid of Hif-1α KO mice demonstrated a decreased phagocytic activity. We are currently investigating whether we could pharmacologically modulate phagocytic activities of alveolar macrophages to regulate the lung metastasis. In conclusion, we demonstrate that HIF-1 in alveolar macrophages is required to protect the irradiated lung from the lung metastasis, possibly in a mechanism involving phagocytosis. Citation Format: Hoibin Jeong, Beom-Ju Hong, Hyung-Seok Choi, Jung-Min Oh, G-One Ahn. Hypoxia-inducible factor-1 in myeloid cells is required to protect the irradiated lung from metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5738.

Myeloid cells (cells that give rise to monocytes and macrophages) are a critical component in the solid tumor microenvironment, promoting angiogenesis and tumor recurrence to therapies. Recent literature have extensively demonstrated that hypoxia-inducible factor (HIF) is a key transcription factor in myeloid cells in responding to hypoxic stimuli including solid tumors and inflammation by secreting various cytokines and growth factors. Although a number of studies have reported that hypoxia exposure to primary macrophages induces HIF-1 activation and subsequent vascular endothelial growth factor (VEGF) production, there had been no mouse model to demonstrate this phenomenon. To better understand the role of transcriptional activation of HIF in pathological macrophages, we have created a new strain of myeloid-specific knockout (KO) mice targeting HIF pathways using hS100A8 as the myeloid promoter. S100A8 is an intracellular calcium binding protein and its expression has been heavily detected in pathological macrophages of many diseases including solid tumors, inflammatory bowel disease, obesity, and rheumatoid arthritis. Upon generating mice deficient for von Hippel Lindau (pVHL) tumor suppressor, the negative regulator of HIF, in myeloid cells, we observed erythema and increased VEGF expression in the bone marrow lysate. Moreover, these mice exhibited an enhanced angiogenesis in the subcutanouesly implanted matrigel plugs, which was accompanied by increased VEGF-VEGFR2 signaling in matrigel. We further found that these phenotypes were dependent on transcriptional activation of HIF-1 as a pharmacological or genetic inhibition of HIF-1α completely suppressed the phenotypes in mice deficient for pVHL in myeloid cells. Importantly, we found that HIF-1 activation in myeloid cells regulate not only VEGF but also S100A8 production and identified that monocytes were the major effector driving angiogenesis. Together these results suggest that transcriptional activation of HIF-1 in myeloid cells plays a critical role in promoting angiogenesis. We are currently investigating how HIF-1 in myeloid cells regulates tumor microenvironment thereby affecting tumor progression. Citation Format: G-One Ahn, Young-Eun Kim, Beom-Ju Hong, Seoyeon Bok, Chan-Ju Lee, Hak Jae Kim, Il Han Kim, Jun Seita, Irving Weissman, J Martin Brown. Hypoxia-inducible factor-1 (HIF-1) in myeloid cells promotes angiogenesis by regulating VEGF and S100A8 production. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr A13. doi:10.1158/1538-7445.CHTME14-A13